Malaria is transmitted by the mature, blood feeding portion of mosquito vector populations. Malaria vector control programs based on indoor residual spraying (IRS) of insecticides are designed to target resting adult Anopheles mosquitoes before or after they have blood fed. When a female mosquito acquires a blood meal, she could also ingest harmful xenobiotics that are present in the blood. During the resting period after feeding, many processes are initiated in order to assist in the digestion and assimilation of the blood. Ultimately, this enables the mosquito to absorb those amino acids needed for the biosynthesis of yolk proteins, which are essential for subsequent egg maturation. Since the regulation of xenobiotic (including insecticides) detoxification enzyme systems is likely to be altered in response to the ingestion of blood, this study aimed to investigate the effect of a blood meal on insecticide tolerance in insecticide resistant and susceptible southern African strains of the major malaria vector Anopheles funestus. Through the use of CDC bottle bioassays it was demonstrated that blood fed An. funestus carrying a pyrethroid resistant phenotype are even more tolerant of pyrethroid intoxication than their unfed counterparts. Using another major malaria vector, An. gambiae, microarray analysis revealed that a general increase in delta class glutathione-s-transferase (GST) expression occured in response to a blood meal. One gene, GSTD3, was over-expressed in both blood fed An. gambiae and An. funestus. Although this gene could not be validated with real time quantitative PCR, it serves as a viable target for future investigations. Since the pyrethroid resistant phenotype of southern African An. funestus has been linked to the over-expression of the duplicate copy gene CYP6P9, the expression levels of both copies of this gene were investigated. CYP6P9 and its copy, CYP6P13, showed a small but significant increase in expression in response to a blood meal. The increased expression of these major effect genes in response to blood feeding may be responsible for the increase in insecticide tolerance seen in the bottle bioassays. In an effort to repeat these experiments on wild caught An. funestus, field material was collected from Karonga in northern Malawi. Specimens were morphologically identified as members of the An. funestus group. However, attempts to molecularly identify them to species level failed. Through the use of ITS2 and D3 sequence analysis, cytogenetics and cross mating studies it was possible to conclude that these wild caught specimens were a new species. They have been provisionally named An. funestus-like.

Anopheles funestus is a major vector of malaria in the southern African region. Insecticide resistance to pyrethroid and carbamate insecticides has been recorded in populations of this species in South Africa and Mozambique. This study aimed to determine the relationship between pyrethroid resistance and gene expression of two closely related genes, CYP6P9 and CYP6P13, by age and sex in a resistant strain An. funestus from southern Africa, FUMOZ-R. The insecticide susceptibility assays showed that percentage survival in both FUMOZ-R sexes significantly decreased as age increased. The mRNA expressions of CYP6P9 and CYP6P13 were higher in FUMOZ-R relative to the insecticide susceptible strain (FANG). The expression of permethrin resistance varies with age in An. funestus FUMOZ-R. The results indicate that other genes may also be involved in insecticide resistance. In addition to this, the expression profile of other metabolic genes involved in insecticide resistance was also investigated. A microarray based approach was used to identify genes differentially expressed in FUMOZ-R and FANG. As the full set of detoxification genes in An. funestus are unknown, this study investigated the utility of the An. gambiae detox chip to screen for differentially expressed detoxification genes in An. funestus. After optimization of the hybridisation conditions, over 90% of the probes showed a positive signal. Only three genes were significantly (P<0.001) differentially expressed in the females, CYP6P9, COI and CYP6M7. The same genes were also significantly differentially expressed in the adult males, together with an additional 21 transcripts. The third part of this study investigated the gene expression in the first instar, fourth instar and 3-day old adults in FUMOZ-R using the An. gambiae detox chip. The variation in metabolic enzyme gene transcription at the different developmental stages in An. funestus are not known. The identification of differentially transcribed genes at the different life stages provides some insight into the role and function of these genes. A large number of cytochrome P450s (monooxygenases), esterases, glutathione S-transferases (GSTs) and other additional genes were differentially expressed in all life stages. This study provided vital information regarding genes potentially involved in pyrethroid resistant and is the first to provide metabolic or detoxifying transcription gene information in An. funestus.

Orientador: Sergio Antonio De Bortoli / Coorientador: Alessandra Marieli Vacari / Banca: Roberto Marchi Goulart / Banca: Raphael de Campos Castilho / Resumo: Um dos principais problemas para o cultivo de brássicas é o dano causado pela traça-das-crucíferas, Plutella xylostella (Linnaeus, 1758) (Lepidoptera: Plutellidae), o que implica na aplicação de grandes volumes de agrotóxicos. Diversos inseticidas são empregados no controle de P. xylostella. Indoxacarbe foi o primeiro inseticida bloqueador de canais de sódio comercializado, com mecanismo de ação diferente do DDT e dos piretroides, apresentando várias vantagens, como baixa toxicidade aos organismos benéficos. Porém, aplicações frequentes de inseticidas para controle de P. xylostella, combinado com fatores biológicos da praga, podem favorecer a seleção de indivíduos resistentes. Até o momento, P. xylostella desenvolveu níveis de resistência a 95 ingredientes ativos, existindo na literatura mais de 55 relatos de resistência para o indoxacarbe, em sete países de diferentes continentes, inclusive no Brasil. Nesse sentido, este estudo objetivou determinar os efeitos subletais e residuais do indoxacarbe em duas populações de P. xylostella, sendo uma coletada no campo e outra de laboratório (suscetível), avaliando e comparando a toxicidade desse inseticida em diferentes concentrações e analisando o comportamento destas duas populações e assim formar uma base teórica para melhorar o manejo desta praga. Foram realizados bioensaios com duas populações (PC- de campo e PL- de laboratório-suscetível) de P. xylostella com o inseticida indoxacarbe na dose comercial (100mg.L-1), avaliando os ... (Resumo completo, clicar acesso eletrônico abaixo) / Abstract: The main problem for brassica crops is the damage caused by the diamondback moth, Plutella xylostella, which has led to the application of large volumes of pesticides. Several insecticides are employed in the control of P. xylostella. Indoxacarbe was the first commercially available sodium channel blocker insecticide, with a different mechanism of action than DDTs and pyrethroids, has several advantages, such as low toxicity to beneficial organisms. However, frequent applications of these insecticides to control P. xylostella, combined with biological factors of the pest are favoring the selection of resistant individuals. So far P. xylostella has developed levels of resistance to 95 active ingredients and there are in the literature more than 55 reports of resistance to indoxacarb in seven countries on different continents, including Brazil. In this sense, the objective was to determine the residual and sublethal effects of indoxacarb in two populations of P. xylostella, one collected in the field and the other in the laboratory (susceptible), evaluating and comparing the toxicity of this insecticide in different concentrations and analyzing the behavior of these two populations and thus form a theoretical basis to improve the management of this pest. Bioassays were performed with two populations (PC- field and PL- laboratory) of P. xylostella with the insecticide in commercial dose indoxacarb (100 mg.L-1), evaluating the effects of residual products 1, 3, 7, 14 and 21 days after spray. LC50 was also estimated and the sublethal effects of this product applied and evaluated after 48 h were evaluated using the sublethal concentrations of LC5 (0.37 and 1.01 mg.L-1), LC15 (0.87 and 2.06 mg. L-1) and LC25 (1.44 and 3.16 mg.L-1) for PL and PC populations by analyzing some biological characteristics of the individuals. The commer... (Complete abstract click electronic access below) / Mestre

No description available.

Spine title: Systemic insecticides in grape vines. Typescript (copy). Includes bibliographical references (leaf. 82-84).

Reported here is an investigation to determine if azinphos-methyl (AZM), an organophosphate insecticide, adducts to hemoglobin, and if so, whether the hemoglobin adduct could be used as a quantitative marker of occupational AZM exposure. We hypothesized that AZM, or a metabolite of AZM, binds to hemoglobin in erythrocytes forming an adducted protein. We administered radiolabled AZM to rats and found a stable, dose-dependent association of radioactivity with hemoglobin. The decline in hemoglobin-associated radioactivity followed the expected kinetics of erythrocyte turnover in rats. We examined hemoglobin isolated from these rats by high-pressure liquid chromatography, liquid scintillation counting, and electrospray ionization mass spectrometry. These analyses provided evidence of AZM or an AZM metabolite binding to one of the beta proteins of hemoglobin. In vitro incubation of AZM with hemoglobin in a liver microsome system indicated an AZM adduct to heme. Further research is necessary to fully characterize the adduct and determine whether this biomarker will be useful for monitoring human exposure to AZM. / Graduation date: 2000

Thesis (Ph. D.)--University of Washington, 2006. / Vita. Includes bibliographical references (leaves 111-119).

Thesis (Ph. D.)--Ohio state University, 1924. / Autobiography. "Reprinted from the Ohio journal of science, vol. XXVIII, no. 1, January, 1928." Bibliography: p. 38.

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